摘要
目的 :了解青藤碱 (sinomenine ,Sin)对PC 12细胞增殖和表达环氧化酶 2 (COX 2 )及合成前列腺素E2(PGE2 )的影响 ,探讨Sin对神经细胞的作用机制。方法 :采用NGF诱导培养PC 12细胞 ,用不同浓度的Sin(0 ,3× 10 -6,30× 10 -6,15 0× 10 -6mol·L-1)预处理后 ,加或不加脂多糖 (LPS) ,分别用3 H TdR法、竞争ELISA、半定量逆转录 聚合酶联反应 (RT PCR)法、细胞酶联免疫法及Western blot法 ,检测PC 12细胞的增殖活性、细胞培养上清液中PGE2 的水平、细胞COX 2mRNA及蛋白表达水平。同时提取各组细胞蛋白 ,测定其核转录因子NF κB活性。结果 :Sin对LPS诱导的PC 12细胞的COX 2mRNA、蛋白表达具有不同程度的抑制作用 ,与Sin浓度呈明显正相关性 ,且对LPS激活的PC 12细胞的核转录因子NF κB活性有明显的抑制作用。实验未观察到Sin对PC 12细胞增殖的影响。结论 :Sin可显著抑制LPS诱导的PC 12细胞COX 2的表达及其产物PGE2 的合成 ,其作用机制可能是通过Sin抑制PC 12细胞核转录因子NF
Objective: To study the effects of sinome ni ne (Sin) on cell proliferation, intracellular expression of cyclooxygenase-2 (C OX-2), and production of PGE 2 in lipopolysaccharide-induced PC-12 cells, To explore the Sin's mechanism on nerve cell. Method: PC- 12 cells were cultured with nerve growing factors (NGF), and pretreated with Sin at various concentrations (0,3×10 -6 ,30×10 -6 ,150×10 -6 mo l·L -1 ) for 2 hours, then with or without stimulation of lipopolysaccharid e (LPS).The proliferation activity of PC-12 cells was determined by 3H -TdR incorporation, and the production of PGE 2 in culture supernatants of PC -12 cells was detected with competitive ELISA. Expression of COX-2 mRNA in PC -12 cells was analyzed by semi-quantitative RT-PCR, and expression of COX-2 protein was estimated by Western blot method and cellular enzyme immunoassay. Nu clear factor-kappa B (NF-κ B) activity in whole-cell extract of PC-12 cells was also measured by an ELISA-based method. Result: The data showed that Sin down-regulated the expression of COX-2 mRN A and protein, and reduced the production of PGE 2 in the LPS-stimulated PC-1 2 cells which correlated with Sin's concentrations positively. In addition, NF- κ B activity in LPS-stimulated cells was suppressed significantly by Sin. No i nhibition of proliferation of PC-12 cells due to Sin treatment was observed. Conclusion: Sin mediates the down-regulation of expressio n of COX-2 and production of induced PGE 2 in PC-12 cells by suppressing the activity of NF-κ B. [
出处
《中国中药杂志》
CAS
CSCD
北大核心
2004年第9期900-903,共4页
China Journal of Chinese Materia Medica
基金
浙江省中医管理局科研基金资助项目 ( 2 0 0 4C0 72 )
